This application is for a laser scanning confocal imaging system to be devoted to time-lapse or real-time imaging of the dynamic state of live cells in several model systems. The instrument will be integrated into an existing bio-imaging facility that will move into newly renovated space within the Dept. of Biological Sciences. The imaging system will be utilized by a core group of major users and will be available to minor users, both within and outside of the Department, on a limited basis (up to 25% time). The three major users (Drs. Dailey, Lilien, and Soll) will use the instrumentation to accomplish goals related to NIH-funded projects. Dr. Dailey will use time-lapse imaging of neurons in rat brain slices transfected with green fluorescent protein (GFP)-fusion protein constructs to study mechanisms of synapse formation and plasticity. He will test the hypothesis that an adhesion molecule, N-cadherin, regulates synapse formation, plasticity, and maintenance. In another project, Dr. Dailey will characterize the motility behaviors of activated microglia, a type of brain-resident cell that plays a major role in clearing dead cells and cellular debris following brain injury. Simultaneous 2- and 3-channel fluorescence time-lapse imaging will be utilized to study the cell-cell interactions between microglia and other cell types in injured brain tissues. Dr. Lilien will use time-lapse imaging of transfected neurons in developing chick neural retina to determine how adhesion molecules affect the dynamic patterns of axon and dendrite growth. These studies will test hypotheses on the functions of distinct protein domains using perturbational agents that affect specific protein-protein interactions important to N-cadherin function and to cross-talk between N-cadherin and b 1-integrins. Dr. Soll will perform analyses on dynamics of cytoskeletal proteins during cell motility and chemotaxis in live Dictyostelium. The dynamic remodeling of GFP-labeled proteins, including actin, myosins I and II, and the Wiscott-Aldrich protein SCAR, will be analyzed in cells reconstructed every 2 sec, using computer-assisted dynamic image analysis systems (2D and 3D DIAS) developed in the Department's W.M. Keck Image Analysis Facility. The technical expertise of the three primary investigators in time-lapse confocal imaging (Dailey), molecular analysis of adhesion molecules and engineering of cDNAs (Lilien), and motion analysis of cells (Soll) provides a high level of synergism between the projects that will assure their success. Together, these studies will contribute to an understanding of molecular mechanisms of cell growth and directed cell movements related to normal tissue development and to functional remodeling under a variety of pathological conditions in humans.